This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. One of the great challenges facing biochemists today is to understand the folding and structural dynamics of proteins. It is clear that the static images of protein structure obtained by X-ray crystallography are often insufficient for mechanistic characterization. Many proteins require complex structural rearrangements for function. Fluorescence spectroscopy allows one to observe protein motions on the ns timescale, but the relatively large size of fluorophores precludes assigning these motions at high resolution. If optical probes could be made sufficiently small, they could provide the time and structural resolution necessary to truly observe and dissect protein motions. To this end, we are developing thioamides, single atom modifications of the peptide backbone, as extremely small fluorescence quenching probes. We have carried out Stern-Volmer quenching experiments of several fluorophores with acetamide or thioacetamide. We would like to use the RLBL pulsed diode laser setup to excite acridone and fluorescein at 408 nm and measure fluorescent lifetimes in the presence of oxoamides and thioamides, both in intermolecular and intramolecular quenching experiments in which the two chromophores are separated by either rigid (Pro) or flexible (Gly/Ser) peptide linkers. These experiments should allow us to confirm the dynamic nature of thioamide quenching.